Even though smoking has been on the decline, there has not been a commensurate decrease in the mortality and adverse effects associated with it, especially in vulnerable populations such as children and minorities (e.g., Hispanic Americans). While the involvement of the well-known first-hand smoke (FHS) and second-hand smoke (SHS) in the pathogenesis of thrombotic diseases is well documented, the contribution of the newly ?discovered? third-hand smoke (THS) form to such disease processes remains unknown. This derives, in part, from: (1) initial lack of knowedlge of THS existence; (2) lack of appreciation for its ?real? negative health consequences; (3) lack of a THS-exosure animal model that mimics real-life scenarios; and (4) lack of studies regarding such consequences on platelet biology. The present application proposes experiments that address fundamental, mechanistic, epigenetic and clinically-relevant translational aspects of the adverse-health effects of the newly ?realized? form of smoking, THS, in the context of thrombotic disease and platelet biology, and in a gender- specific manner. Studies are also proposed to investigate, in a similar fashion, the toxicants that underlie THS effects on platelets and associated diseases. These studies are of paramount significance given the skepticism regarding THS existence and/or the fact that its dangers are/remain underestimated/unappreciated, despite evidence that it is more toxic than SHS. The Aims of our proposal are: Aim 1. Investigate the impact of THS-exposure on platelet-dependent disease states. While compelling recent evidence revealed that exposure to THS does exert negative health effects, its impact on platelet- dependent diseases and the contribution of gender to these effects are still unknown. To address this issue, we will determine the ramifications of THS exposure on normal hemostasis and platelet counts, in a dose-, time-, and gender-dependent fashion. Subsequent studies will examine whether THS participates in the development of thrombotic disease, as well as investigate the role of the coagulation system in mediating its toxicity. Our preliminary data shows, for the first time, that THS modulates physiological hemostasis, suggesting that it may increase the risk of cardiovascular disease. Aim 2. Investigate the mechanism by which THS-exposure modulates platelet function. While our preliminary studies revealed that THS modulates hemostasis, the mechanism, by which it modules platelet function remains to be investigated. Thus, the overall goal of the experiments proposed in this section is to determine the significance of THS-exposure on the various platelet functional responses, cytokines, platelet epigenetics, thrombosis ?markers? and other blood cells. Studies are also proposed to investigate whether THS effects are receptor mediated. It is noteworthy that our preliminary data shows that THS does enhance platelet function (e.g., aggregation). Aim 3. Define THS toxins with potent impact on platelet-dependent disease and function. While some progress has been made regarding specific FHS and SHS toxicants/ingredients that impact platelet activation, virtually nothing is known in the context of THS. To this end, recent studies, including those by the California THS Consortium (for which Dr. Martins-Green was a founding member), have shown that nicotine, cotinine, 3- ethynylpyridine, benzo(a)pyrene, NNAL, as well as acrolein, are amongst the major THS toxicants. Thus, the impact of these toxicants, alone or in combination, on platelet function and related disease will be investigated, as per the approaches described in Aims 1 and 2; in order to identify the most potent toxicants. To this end, our preliminary data revealed that cotinine- one of the major components of THS- does indeed enhance platelet function. Collectively, these experiments will make significant contributions to our understanding of the consequences of THS exposure (an unappreciated health threat) on platelet activation and cardiovascular human health, as well as the mechanism (e.g., epigenetics) and toxicants by/through which it exerts these effects, in a gender-specific manner.